Transforming growth factor-beta increases breast cancer stem cell population partially through upregulating PMEPA1 expression.

The prostate transmembrane protein, androgen-induced 1 (PMEPA1) has been previously shown to promote solid malignancies in a variety of cancers, but the role and mechanisms of PMEPA1 in breast cancer has not been fully addressed. Here, we found that PMEPA1 was upregulated in breast cancer cell lines as well as in a set of clinical invasive breast ductal carcinomas. Interestingly, depletion of PMEPA1 decreased breast cancer stem cell (CSC)-enriched populations, while ectopic overexpression of PMEPA1 increased breast CSC-enriched populations. Furthermore, transforming growth factor-ß (TGF-ß) treatment was also found to upregulate PMEPA1 expression and the CSC-enriched populations in triple-negative breast cancer cell lines. TGF-ß-induced PMEPA1 expression partially contributed to TGF-ß-induced breast CSC maintenance. These findings suggest that TGF-ß-PMEPA1 axis might provide new diagnosis and therapeutic targets for breast cancer treatment.

A short report: PAMM, a novel antioxidant protein, induced by oxidative stress.

Reactive oxygen species (ROS) play a central role in estrogen deficiency-induced bone loss. We previously identified and characterized a novel member of the Peroxiredoxin (PRX) like 2 family that we called PAMM: Peroxiredoxin Activated in M-CSF stimulated Monocytes, a redox regulatory protein that modulates osteoclast differentiation in vitro. In this study, we report increased PAMM expression in H2O2-treated cells and in bones from ovariectomized (OVX) mice 4 weeks after surgery, models for oxidative stress in vitro and in vivo, respectively. We also detected increased PAMM abundance and phosphorylated Akt in OVX mice treated with estrogen. In addition, Wortmannin, a specific PI3Kinase inhibitor and Rapamycin, an inhibitor of the PI3Kinase/Akt pathway, blocked Akt phosphorylation and stimulation of PAMM expression by M-CSF. These results indicate that M-CSF-induced PAMM expression is mediated by Akt phosphorylation. Our data also suggest that estrogen-induced PAMM expression is mediated by phosphorylation of Akt. These findings point to PAMM as a potential candidate for Akt-mediated protection against oxidative stress.

[Differential expression of miRNAs in myocardial tissues of rats with lipopolysaccharide- induced endotoxemia].

OBJECTIVE: To analyze the differential miRNA expression profile in the myocardium of rats with lipopolysaccharide (LPS)-induced endotoxemia and explore the role of miRNA in endotoxin-induced myocardial injury. METHODS: Twenty male SD rats received intraperitoneal injection of 10 mg/kg LPS (n=10) or an equivalent amount of saline solution (n=10). At 24 h after LPS injection, the rats were sacrificed to detect myocardial expressions of TLR4, TNF-α and IL-1ß using real-time PCR and for observing myocardial ultrastructures under transmission electron microscopy. The differentially expressed miRNA in the myocardium were detected using a miRNA array, and the common differentially expressed miRNAs were selected for verifying their actual expressions using real-time PCR. RESULTS: TLR4, TNF-α and IL-1ß were over-activated in the myocardium of LPS-treated rats, in which mitochondria swelling, structural damaged and cytoplasmic vacuoles were observed. In LPS-challenged rats, miR-194-3p, miR-344a-3p, miR-465-3p, miR-501-5p, miR-3596c, miR-185-3p, and miR-877 were found up-regulated significantly, whereas miR-208b-3p, miR-547-3p, miR-141-3p, miR-28-5p, and miR-3585-5p down-regulated in the myocardium. CONCLUSION: Significant differential expression of the miRNAs occurs in the myocardium of LPS-treated rats, suggesting their involvement in endotoxin-induced myocardial injury.

The comparison of methods to identify the presence of fibrocytes in formalin-fixed, paraffin-embedded archival cardiac tissue with coronary heart disease.

The purpose of this study was to find the optimal technical approach to identify the presence of fibrocytes in formalin-fixed, paraffin-embedded archival cardiac tissue with CHD (coronary heart disease). Using the coexpression markers CD45 and αSMA, the presence of fibrocytes was examined by three different methods, including double immunohistochemistry staining, combination labeling of immunohistochemistry and immunofluorescence and double immunofluorescence labeling. Double immunohistochemistry staining was very difficult to identify the CD45(+)/αSMA(+) fibrocytes. Although combination staining of immunohistochemistry and immunofluorescence has made it possible to evaluate the co-localization of CD45 and αSMA in the fibrocytes, this method was prone to produce many false positive cells. In contrast, CD45(+)/αSMA(+) fibrocytes could be clearly recognized by double immunofluorescence labeling. In conclusion, double immunofluorescence labeling is the optimal technical approach to identify the presence of fibrocytes in routinely processed cardiac tissue with CHD.

[Expression of PEPT2 mRNA in lung tissue of rats with pulmonary fibrosis].

BACKGROUND AND OBJECTIVE: Pulmonary fibrosis is a common pathological phenomenon in lung cancer patients after chemotherapy or radiotherapy. It is also a key hindrance to the transport of drugs to lung tissue. Peptide transporters have become a target of the rational design of peptides and peptide drugs. The aim of this study is to investigates the expression of peptide transporter 2 (PEPT2) mRNA in the lungs of rats with bleomycin (BLM)-induced pulmonary fibrosis. METHODS: Fifty healthy adult Sprague-Dawley rats were randomly divided into five groups. One group was untreated (control), the second group was injected with normal saline solution (NS), and the three remaining groups were treated with a single dose of bleomycin to induce pulmonary fibrosis (BLM). Rats from the NS group were killed by exsanguination on day 14. Rats from the BLM group were killed by exsanguination on days 7, 14, and 28. The lung samples were observed under light microscopy and the hydroxyproline concentration was determined. The expression levels of PEPT2 mRNA were measured by RT-PCR. RESULTS: The morphological study showed that collagenous fiber proliferated in the lungs of rats injected with BLM, indicating pulmonary fibrosis. This proliferation was apparent at 14 d post-injection and especially at 28 d post-injection. Hydroxyproline levels increased seven days post-injection compared with the control group and NS group, but there was no significant statistical difference (P>0.05). Hydroxyproline levels significantly increased (P<0.05) 14 d and 28 d post-infection. The change in the lung tissue pathology coincided with the change in hydroxyproline levels. There were no significant changes of pulmonary PEPT2 mRNA expression levels among the different groups (P>0.05). CONCLUSION: PEPT2 is a potential peptide drug target in the treatment of pulmonary fibrosis, although there were no significant changes of PEPT2 mRNA expression in the lungs of rats with bleomycin-induced pulmonary fibrosis.

Extracorporeal cardiac shock wave therapy ameliorates myocardial fibrosis by decreasing the amount of fibrocytes after acute myocardial infarction in pigs.

OBJECTIVES: Extracorporeal shock wave (SW) therapy ameliorates cardiac remodeling after acute myocardial infarction (AMI). However, it remains to be examined whether and how SW therapy ameliorates myocardial fibrosis after AMI. Fibrocytes are associated with myocardial fibrosis. Thus, we examined whether SW therapy ameliorates myocardial fibrosis and whether fibrocytes are associated after AMI in pigs. MATERIALS AND METHODS: AMI was created by coronary embolism. Twenty-five pigs were divided into three groups: AMI+SW group (AMI with SW therapy, n=15), AMI group (without SW therapy, n=5), and sham+SW group (SW therapy without AMI, n=5). The collagen area fraction was examined by Masson's trichrome staining. The presence of fibrocytes was identified by immunofluorescence and confocal microscopy. The location of CXCL12 was examined by immunohistochemistry. RESULTS: Compared with the AMI group, the AMI+SW group showed significantly ameliorated myocardial fibrosis in terms of collagen area fraction (27.21±8.13 vs. 10.13±4.96, P<0.05) and reduced fibrocytes (CD34/α-smooth muscle actin: 35.40±11.72 vs. 12.27±7.71, P<0.05; CXCR4/α-smooth muscle actin: 40.80±8.96 vs. 16.54±6.38, P<0.05). There were positive correlations between the collagen area fraction and the number of fibrocytes (r=0.936; P<0.05) and between the number of CXCR4 fibrocytes and the SDF-1/CXCL12 cells (r=0.802; P<0.05) in the three groups. CONCLUSION: The results show that SW therapy ameliorates myocardial fibrosis after AMI in pigs, which is associated with the decreased amount of fibrocytes.

Effects of sevoflurane on pulmonary cytosolic phospholipase A2 and clara cell secretory protein expressions in rabbits with one-lung ventilation-induced lung injury.

OBJECTIVE: To investigate the effects of sevoflurane on cytosolic phospholipase A2 (C-PLA2) and clara cell secretory protein (CCSP) in lung tissues of rabbits with one-lung ventilation (OLV)-induced lung injuries. METHODS: Thirty-six healthy Japanese white rabbits were randomized into sham-operated group, OLV group, and OLV plus sevoflurane group subdivided into 4 subgroups with sevoflurane concentrations of 1%, 2%, 3% and 4%. CCSP and C-PLA2 mRNA and protein expressions in rabbit lung tissues were detected by Western blotting and real-time PCR, and the content of arachidonic acid (AA) was measured using ELISA. The severities of the lung injury were evaluated according to lung wet/dry weight (W/D) ratio and histological scores. RESULTS: In the OLV group and OLV+ sevoflurane groups, pulmonary CCSP expressions were significantly lower, while C-PLA2 expression, lung W/D ratios and lung histological scores were significantly higher than those in the sham-operated group (P<0.05). Compared with OLV group, the OLV+sevoflurane groups showed significantly increased expressions of CCSP and reduced C-PLA2 expression, lung W/D ratios and histological scores (P<0.05). In the 4 OLV+sevoflurane groups, CCSP expressions underwent no significant changes as sevoflurane concentration increased, but C-PLA2 expressions, lung W/D ratios and histological scores all decreased gradually as the concentrations of sevoflurane increased (P<0.05). CONCLUSION: OLV can result in down-regulated CCSP expressions and up-regulated C-PLA2 expressions in rabbit lung tissues. Sevoflurane can protect against OLV-induced acute lung injury possibly by inhibiting C-PLA2 expression via up-regulation of CCSP expressions or through other mechanisms resulting in down-regulated expression of C-PLA2.

[Relationship between adipose expression of 3-phosphoinositide-dependent protein kinase 1 and glycometabolism in a mouse model of hyperhomocysteinemia].

OBJECTIVE: To study the effect of homcysteine on the expression of 3-phosphoinositide-dependent protein kinase 1 (PDK1) in the adipose tissue and explore whether PDK1 inhibits p-Akt(Thr-308) expression and affect PI3K/Akt signal pathway to decrease glucose uptake and utilization. METHODS: Forty mice were randomly divided into 4 groups (n=10), namely the fasting control group, feeding control group, fasting hyperhomocysteinemia group, and feeding hyperhomocysteinemia group. In the two hyperhomocysteinemia groups, the mice were given water containing 1.5% methionine to induce hyperhomocysteinemia. Blood glucose and insulin levels in each group were determined, and the expressions of PDK1 and Akt mRNA in the adipose tissue were detected using RT-PCR; the expressions of PDK1, p-Akt(Thr-308) and Akt proteins were detected using Western blotting. RESULTS: In the fasting and feeding hyperhomocysteinemia groups, blood glucose and insulin levels were significantly higher than those in the two control groups. The expressions of PDK1 mRNA and PDK1 and p-Akt(Thr-308) proteins were reduced in the two hyperhomocysteinemia groups, but Akt mRNA and protein expressions were comparable with those in the control groups. CONCLUSION: Homocysteine lowers the uptake and utilization of glucose by down-regulating PDK1 expression and affecting PI3K/Akt signal pathway to cause disturbance of glucose metabolism.

Protective mechanisms of sevoflurane against one-lung ventilation-induced acute lung injury: role of cyclooxygenase-2 and 5-lipoxygenase pathways.

OBJECTIVE: To explore the protective mechanisms of sevoflurane against acute lung injury (ALI) induced by one-lung ventilation (OLV) in view of cyclooxygenase-2 (COX2) and 5-lipoxygenase (5-LOX) pathways. METHOD: Eighteen healthy Japanese white rabbits were randomized into sham-operated group (S group), OLV group (O group) and OLV + sevoflurane group (OS group). COX2 and 5-LOX protein and mRNA expressions in the lungs were detected by Western blotting and real-time PCR, respectively. Prostaglandin I2 (PGI2), thromboxane A2 (TXA2) and leukotrienes B2 (LTB2) in the lung tissues were quantified with ELISA. Histological scores and lung wet/dry weight (W/D) ratios were determined for lung injury assessment. RESULTS: COX2 and 5-LOX protein and mRNA expressions and the contents of LTB2, TXA2 and PGI2 in the lungs, lung W/D ratio and histological scores were significantly higher while PGI2/TXA2 ratio was significantly lower in O group and OS group than in S group (P<0.05). Compared with those in O group, COX2 and 5-LOX expressions, pulmonary contents of LTB2, TXA2 and PGI2, and lung W/D ratio all decreased significantly but PGI2/TXA2 ratio was significantly elevated in OS group (P<0.05). CONCLUSION: OLV may activate COX2 and 5-LOX pathways to result in increased production of arachidonic acid metabolites. Sevoflurane protects against OLV-induced ALI probably by reducing AA metabolites and regulating PGI2/TXA2 ratio through inhibitions of COX2 and 5-LOX pathways.

siRNA-Act1 inhibits the function of IL-17 on lung fibroblasts via the NF-κB pathway.

BACKGROUND: Interleukin (IL)-17-producing T lymphocytes play a role in pulmonary fibrosis, but the possible mechanism of IL-17 on lung fibroblasts remains uncertain. OBJECTIVES: To explore the role and possible mechanism of IL-17 on lung fibroblasts. METHODS: A mouse model of pulmonary fibrosis was established by intratracheal administration of 5 mg/kg bleomycin. At 14 days following bleomycin administration the pulmonary fibroblasts were isolated, cultured and identified. siRNA for activator 1 (Act1) were transfected into lung fibroblasts, which were cocultured with IL-17. The NF-κB pathway was detected for IL-17 on the lung fibroblasts. RESULTS: IL-17R was increased significantly at 14 days in the bleomycin-induced pulmonary fibroblast model, exogenous IL-17 significantly promoted the proliferation of the pulmonary fibroblasts in primary culture and obviously increased the expression of α-smooth muscle actin and type I and type III collagen in the fibroblasts. We found that IL-17 rapidly activated the NF-κB signaling pathway through activated phosphorylated p65 and IκB, and all roles of IL-17 on lung fibroblasts were inhibited under the interference for the expression of Act1 in lung fibroblasts. CONCLUSION: IL-17 may directly promote the proliferation, transformation and collagen synthesis of lung fibroblasts via the NF-kB signaling pathway, which can be inhibited by the interference for the expression of Act1.

Fibrocytes are associated with the fibrosis of coronary heart disease.

Fibrocytes contribute significantly to fibrosis in many cardiac diseases. However, it is not clear whether fibrocytes are associated with the fibrosis in coronary heart disease (CHD). The aim of this study was to determine whether fibrocytes are involved in cardiac fibrosis in CHD. We identified the presence of fibrocytes in CHD heart by immunofluorescence and confocal microscopy, examined the collagen volume fraction by Masson's Trichrome staining, and evaluated the correlation between fibrocytes and cardiac fibrosis. In conjunction, we examined the location of CXCL12, a homing factor and specific ligand for CXCR4, by immunohistochemistry. Fibrocytes were identified in 26 out of 27 CHD hearts and in 10 out of 11 normal hearts. Combinations, including CD34/αSMA, CD34/procollagen-I, CD45/αSMA, CXCR4/procollagen-I and CXCR4/αSMA, stained significantly more fibrocytes in CHD hearts as compared with those in normal hearts (p<0.05). There were positive correlations between the collagen volume fraction and the amount of fibrocytes (r=0.558; p=0.003<0.01) and between the number of CXCR4(+) fibrocytes and the CXCL12(+) cells (r=0.741; p=0.000<0.01) in CHD hearts. Based upon these findings, we conclude that fibrocytes, likely recruited through the CXCR4/CXCL12 axis, may contribute to the increase in the fibroblast population in CHD heart.

Therapeutic effect of intravenous bone marrow-derived mesenchymal stem cell transplantation on early-stage LPS-induced acute lung injury in mice.

OBJECTIVE: To investigate the effect of intravenous bone marrow-derived mesenchymal stem cell (MSC) transplantation for early intervention of lipopolysaccharide (LPS)-induced acute lung injury (ALI) in mice. METHODS: Thirty-six mice were randomized into control group, PBS-treated ALI group, and MSC-treated ALI group. In the latter two groups, mouse models of ALI were established by intranasal instillation of LPS, and 1 h later, the 4th passage of MSCs isolated from the bone marrow of mice or PBS were administered via the tail vein. The histological findings, lung wet/dry (W/D) weight ratio, neutrophil count and protein and cytokine contents in the bronchoalveolar lavage fluid (BALF), and myeloperoxidase (MPO) level in the lung tissue were analyzed at 24 h after MSC administration. Engraftment of MSCs in the recipient lung was determined by fluorescent PKH26 staining and flow cytometry. RESULTS: Compared with the control group, PBS-treated ALI group showed significantly higher protein levels, tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and neutrophil count in the BALF and MPO content in the lung tissue, with also severe damage of lung histology. MSCs administration significantly reduced the lung W/D weight ratio, the levels of protein, TNF-α, IL-6 and neutrophil count in the BALF and MPO content in the lung tissue, and obviously lessened the lung injury 24 h after the transplantation. MSC administration also significantly increased the level of IL-10 in the BALF. CONCLUSION: Intravenous MSC transplantation can effectively improve the lung histology, attenuate the inflammatory response, reduce pulmonary edema in the early stage of LPS-induced ALI in mice, and such effects are independent of MSC engraftment in the lungs.

[Effects of interleukin-17 on murine pulmonary fibroblast proliferation, transformation and collagen synthesis].

OBJECTIVE: To investigate the effects of interleukin-17 (IL-17) on the proliferation, transformation and collagen synthesis of the lung fibroblasts in mice with bleomycin-induced pulmonary fibrosis. METHODS: In a mouse model of pulmonary fibrosis established by intratracheal administration of 5 mg/kg bleomycin, the dynamic expressions of IL-17/IL-17 receptor (IL-17R) mRNAs were detected by RT-PCR. At 14 days following bleomycin administration, the pulmonary fibroblasts were isolated, cultured and identified. MTT assay was used to assess the proliferation of the pulmonary fibroblasts in response to IL-17 treatment at different concentrations, and RT-PCR and Western blotting were employed to examine the mRNA and protein expressions of α-smooth muscle actin (α-SMA) and types I and III collagen. RESULTS: IL-17/IL-17R mRNA levels were increased obviously in the pulmonary fibroblasts of rats with pulmonary fibrosis, and the highest expressions occurred at 14 days following bleomycin administration. Exogenous IL-17, at the optimal concentration of 50 ng/ml, significantly promoted the proliferation of the pulmonary fibroblasts in primary culture and obviously increased α-SMA expression and types I and III collagen synthesis in the fibroblasts. CONCLUSION: IL-17 can promote the proliferation, transformation, and collagen synthesis of the pulmonary fibroblasts from rats with bleomycin-induced pulmonary fibrosis.

The role of all-trans retinoic acid in bleomycin-induced pulmonary fibrosis in mice.

Much evidence suggests that immune imbalance in the lung plays a crucial role in the development of pulmonary fibrosis. Recently, all-trans retinoic acid (ATRA) shifting the regulatory T/T-helper 17 (Treg/Th17) profile had been proven in some diseases. However, to date, the effect of ARTA of pulmonary fibrosis has not been examined from this aspect. The objective of this study was to study the effect of ATRA on bleomycin-induced pulmonary fibrosis in mice and its possible mechanism. Pulmonary fibrosis was induced in C57BL/6 male mice by intratracheal instillation of bleomycin (5 mg.kg(-1)), which were randomly divided into control, bleomycin, and ATRA groups. Five mice in each group were sacrificed on day 28 after intratracheal instillation. Hemotoxylin and eosin (H&E) and Masson staining were used for pathological examination, and hydroxyproline in lung tissue was measured. Interleukin (IL)-17A protein expression was observed in lung with immunohistochemistry. The expression of IL-17A, IL-10, IL-6, and transforming growth factor (TGF)-ß mRNAs were detected by reverse transcriptase-polymerase chain reaction (RT-PCR). Th17 and Treg expression in spleen lymphocytes were measured by flow cytometry. H&E and masson staining and expression of hydroxyproline showed that ATRA significantly alleviated lung fibrosis than in the bleomycin group. The expression of IL-17A, IL-10, IL-6, and TGF-ß mRNAs were higher in the bleomycin group than in the normal group. ATRA can decrease these cytokines except for IL-10. CD4+CD25+ Treg cell ratio in the bleomycin group was significantly lower than normal, but CD4+IL-17+ T cells was higher; ARTA reversed this kind of expression. ATRA may ease the bleomycin-induced pulmonary fibrosis by inhibiting the expression of IL-6 and TGF-ß, shifting the Treg/Th17 ratio and reducing the secretion of IL-17A.

Role of asymmetric dimethylarginine in acute lung injury induced by cerebral ischemia/reperfusion injury in rats.

OBJECTIVE: To determine the role of asymmetric dimethylarginine (ADMA) in acute lung injury induced by cerebral ischemia/reperfusion (I/R) injury in rats. METHODS: Adult male SD rats were randomly divided into 4 groups, namely the sham-operated group (S), cerebral I/R model group, ADMA+I/R group, and dimethylarginine dimethylaminohydrolase (DDAH)+I/R group. In the latter 3 groups, acute lung injury was induced by left middle cerebral artery occlusion for 120 min. After a 24-h reperfusion, the rats were sacrificed and the activities of nitric oxide synthase (NOS) and contents of nitric oxide (NO) were measured using reductase and colorimetric assay. The mRNA and protein expressions of protein kinase C (PKC) and myosin light chain kinase (MLCK) in the lung tissues were detected with RT-PCR and Western blotting, respectively. The contents of ADMA in the bronchoalveolar lavage fluid (BALF) and blood flowing into and out of the lungs were measured by ELISA. RESULTS: Cerebral I/R injury caused significantly elevated ADMA levels in the BALF and blood flowing into the lungs, and obviously lowered the NO concentration and NOS activity in the lung tissues (P<0.05). Following cerebral I/R injury, MLCK and PKC mRNA and protein expressions were significantly upregualted in the lung tissues (P<0.05). Exogenous DDAH obviously decreased the levels of ADMA in the BALF and blood flowing into the lungs, increased NO concentration and NOS activity, and down-regulated MLCK and PKC mRNA and protein expressions in lung tissues of rats with cerebral I/R injury (P<0.05). CONCLUSION: ADMA contributes to the development of acute lung injury following cerebral I/R injury in rats by upregulating MLCK and PKC expression. ADMA may serve as a novel therapeutic biomarker and a potential therapeutic target for acute lung injury induced by cerebral I/R injury.

Paragonimus worm from a New Guinea native in 1926.

OBJECTIVE: To reobserve and research the specimen of Paragonimus worm found in the left lung of a New Guinea native in 1926, which was previously identified as Paragonimus westermani Kerbert or Paragonimus ringeri Cobbold. METHODS: Using reconstructive software and microscopy to observe some organs of the worm, and compared with other species of paragonimus. RESULTS: The three dimensional (3D) views of ovary and two testes of New Guinea specimen showed that the ovary was clearly divided into six lobes. These two testes were situated oppositely in the body. One teste was divided into four branches, while another was divided into five. The cuticular spines were arranged in groups over the entire skin covered in a slide, each group was consisted of two to four single spine. CONCLUSIONS: Based on 3D views and measurements, we reclassified it as Paragonimus siamensis. This was also the first report of human case infected by Paragonimus siamensis.

Effect of asymmetric dimethylarginine on MIF expression and TNF-α and IL-8 secretion in THP-1 monocytes-derived macrophages.

OBJECTIVE: To investigate the effect of ADMA on macrophage migration inhibitory factor (MIF) expression and tumor necrosis factor-α (TNF-α) and IL-8 secretion in THP-1 monocyte-derived macrophages. METHIDS: THP-1 monocytes were induced to differentiate into macrophages by a 24-h incubation with 160 nmol/L PMA. The THP-1 monocyte-derived macrophages were exposed to different concentrations of ADMA for 24 h, and the changes in MIF mRNA and protein expressions were analyzed with RT-PCR and Western blotting, respectively. Enzyme-linked immunosorbent assay was used to detect the levels of TNF-α and IL-8 in the supernatant of THP-1-derived macrophages following ADMA treatments. RESULTS: ADMA obviously up-regulated MIF mRNA and protein expressions in THP-1-derived macrophages in a concentration- dependent manner. Exposure of the cells to 15 µmol/L ADMA for 24 h showed the most potent effect in up-regulating MIF mRNA and protein expressions. ADMA treatment also resulted in a dose-dependent increase of the levels of TNF-α and IL-8 in the culture supernatant of the macrophages, and the peak levels occurred following the treatment with 15 µmol/L ADMA. CONCLUSION: ADMA can up-regulate MIF expression and induce TNF-α and IL-8 secretion in THP-1 monocyte-derived macrophages.

Asymmetric dimethylarginine upregulates the expression of ACAT-1 in THP-1 macrophage-derived foam cells.

OBJECTIVE: To investigate the effects of asymmetric dimethylarginine (ADMA) on ACAT-1 expression and cholesterol content in THP-1-derived macrophages and foam cells. METHODS: THP-1 cells were induced to differentiate into macrophages and further into foam cells. The macrophages and foam cells were exposed to different concentrations (0, 3.75, 7.5, 15, and 30 µmol/L) of ADMA for varying time lengths (6, 12, and 24 h), and the changes in ACAT-1 mRNA and protein levels in the cells were measured with RT-PCR and Western blotting. The cellular cholesterol content was measured with enzyme-linked colorimetry assay. RESULTS: In THP-1-derived macrophages and foam cells, the expression levels of ACAT-1 mRNA and protein and cellular cholesterol content increased significantly in response to ADMA treatment in a time- and concentration-dependent manner. CONCLUSION: ADMA may play an important role in inducing foam cell formation from macrophages. ACAT-1 inhibition targeting the macrophages and foam cells may serve as a potential therapeutic target in the treatment of atherosclerosis.

[Change of cytochrome c in postconditioning attenuating ischemia-reperfusion-induced mucosal apoptosis in rat intestine].

The present study aimed to investigate the change of cytochrome c in postconditioning-attenuated ischemia-reperfusion (I/R)-induced mucosal apoptosis in rat intestine compared with ischemic preconditioning (IPC). Using rat model of intestine I/R injury, male Sprague-Dawley rats weighing 220-250 g were divided into 4 groups which were Sham operation group, I/R group, IPC group and ischemic postconditioning (IPOST) group. In these groups, I/R procedure was performed by the occlusion of the superior mesenteric artery (SMA) for 45 min followed by reperfusion for 1 h. In Sham group, there was no intervention. In IPC group, SMA was occluded for 5 min and reperfused for 5 min, for two cycles, before the prolonged occlusion. In IPOST group, three cycles of 30-s reperfusion and 30-s reocclusion were preceded at the start of reperfusion. After the reperfusion, the small intestines were sampled for experimental detection. Intestinal mucosal mitochondrial membrane potential was detected by confocal laser scanning microscopy. Expressions of cytochrome c and caspase-3 proteins were detected using Western-blot method. The apoptosis of intestinal mucosal cells was determined with agarose gel electrophoresis and deoxynucleotidyl transferase mediated dUTP-biotin nick-end labeling (TUNEL) technique. Compared with I/R group, the mitochondrial membrane potentials and the expressions of cytochrome c protein were significantly increased, while the expressions of caspase-3 and the apoptotic rates were decreased in IPOST and IPC groups (P<0.05). There were no significant differences between IPOST and IPC groups (P>0.05). These data provide substantial evidence that IPOST attenuates I/R-induced mucosal apoptosis by reducing the release of cytochrome c from mitochondria in the rat small intestine.